DNA, isolated from temperate coliphage lambda h80dlacps and containing the sequence coding for beta-galactosidase, was used to initiate in vitro synthesis of the enzyme in the presence and absence of various carcinogens, especially alkylating agents. Beta-Galactosidase activity was measured colorimetrically by hydrolysis of O-nitrophenyl-beta-D-galactopyranoside. Inhibition of synthesis of enzyme activity resulting from carcinogen interaction with DNA and with other components by the system is measured. Inhibition of beta-galactosidase synthesis by methyl- and ethylnitrosourea is strictly a consequence of alkylation, and is equivalent at equal levels of total alkylation. This is inconsistent with hypotheses postulating that certain types of alkylation reaction such as 7-alkylguanine formation are insignificant. Carbamylation was not an important reaction with these compounds and, although demonstrable, did not contribute to inhibition of DNA function by methylnitrosourea.